Conventionally, somatic homologous recombination is thought of as one factor that produces genetic diversity. For example, in a chicken-derived B cell lines, it is known that DNA recombination occurs at immunoglobulin locus for antibodies (Buerstedde et al., EMBO J. (1990) 9:921-927). Creating various protein factors using such DNA recombination is thinkable, but in actuality, the recombination frequency is extremely low, so it was difficult to use by itself for the creation of protein factors.
Additionally, when producing antibodies, animals such as rabbits and mice are normally used, but in cases where it is desired for even more diverse antibodies to be obtained, it was necessary to immunize a large number of individual animals with the antigens. Additionally, the titers of antibodies obtained depended on differences between individual animals, and the properties of the antigens, so that it was difficult to stably obtain diverse antibodies at high titers.
Consequently, obtaining antibodies by using DNA homologous recombination that occurs in the aforementioned chicken-derived B cell lines and the like was conceived of, and it is thought that in principle, it would become possible to synthesize diverse antibodies with this method. However, as described above, since the DNA homologous recombination frequency is extremely low, it was thought that it was extremely difficult in reality to artificially create antibodies for specific antigens with cultured cells.
On the other hand, in recent years, in XRCC2 and XRCC3 knockout DT40 cell lines (chicken-derived B cell lines), it has been reported that the frequency of somatic cell mutation increases (Sale et al., Nature (2001) 412:921:926). The use of such somatic cell mutations has been considered, and in actuality, attempts have been made to increase antibody affinity to antigens by using Ramos cells and XRCC2 and XRCC3 knockout DT40 cells. However, antibodies obtained using such methods show no specificity, and bind to various proteins (Cumbers et al., Nat. Biotechnol. (2002) 20 (11):1129-1134). Since somatic cell mutations have a secondary use in vivo for affinity maturation, it is thought that making large-scale changes in antibody specificity would be difficult.